Coumarin-30 Enables Site-Resolved Detection of Tubulin Ligands by Microscale Thermophoresis
Anisimov M.N. Boichenko M.A. Sivachev A.A. Romanov A.N. Lifshits I.A. Borzunova J.N. Janibekova M. Mustyatsa V.V. Tikhonov T.P. Plodukhin A.Y. Shorokhov V.V. Tafeenko V.A. Ratmanova N.K. Zhokhov S.S. Novikov R.A. Kechko O.I. Makarov A.A. Mitkevich V.A. Andreev I.A. Vorobjev I.A. Trushkov I.V. Ivanova O.A. Gudimchuk N.B.
16 March 2026John Wiley and Sons Inc
Angewandte Chemie - International Edition
2026#65Issue 12
Tubulins are among the most successful targets for cancer chemotherapy. However, the emergence of drug resistance stimulates the continuous search for novel chemotherapeutics. We discovered that coumarin-30, a widely available laser dye, binds to the colchicine site of tubulin, inhibiting microtubule dynamics and cancer cell division at submicromolar concentrations. Exploiting the excellent fluorescent properties of coumarin-30, we developed a fast, accurate, and cost-effective coumarin-30-based microscale thermophoresis (C-MST) assay as an express method for detecting tubulin–ligand interactions and discriminating colchicine site binders from ligands targeting other protein pockets. Using this assay, we identified several potent tubulin polymerization inhibitors associating with the colchicine site and validated them through in vitro microtubule dynamics and cell cycle assays in cancer cells. Furthermore, the C-MST assay was demonstrated to detect ligands targeting a novel binding site on tubulin, recently established through crystallographic fragment screening. We confirmed detection of a small-molecule ligand targeting that site and further designed and characterized a series of its analogs. The ability of the C-MST assay to detect tubulin binders regardless of their binding site or their effect on microtubule dynamics opens new avenues for developing unconventional modulators of tubulin–tubulin and tubulin–effector interactions thereby facilitating anticancer drug discovery.
Coumarin-30 , Drug discovery , High-throughput screening , Microscale thermophoresis , Tubulin
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Department of Physics, M. V. Lomonosov Moscow State University, Leninskie Gory 1–2, Moscow, 119991, Russian Federation
Center for Theoretical Problems of Physicochemical Pharmacology, Russian Academy of Sciences, Srednyaa Kalitnikovskaya 30, Moscow, 109029, Russian Federation
Department of Chemistry, M. V. Lomonosov Moscow State University, Leninskie Gory 1–3, Moscow, 119991, Russian Federation
N. N. Semenov Institute of Chemical Physics, Russian Academy of Sciences, Kosygina 4, Moscow, 119991, Russian Federation
National Laboratory Astana, Kabanbai batyr 53, Astana, 010000, Kazakhstan
N. D. Zelinsky Institute of Organic Chemistry, Russian Academy of Sciences, Moscow, 119991, Russian Federation
Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilova 32, Moscow, 119991, Russian Federation
Department of Biology, School of Sciences and Humanities, Nazarbayev University, Kabanbai batyr 53, Astana, 010000, Kazakhstan
Department of Physics
Center for Theoretical Problems of Physicochemical Pharmacology
Department of Chemistry
N. N. Semenov Institute of Chemical Physics
National Laboratory Astana
N. D. Zelinsky Institute of Organic Chemistry
Engelhardt Institute of Molecular Biology
Department of Biology
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